Control of Microbial Growth

A few terms Bacteriostatic: inhibits bacterial growth Bactericidal: something capable of killing bacteria Antiseptic: an agent that is used to inhibit/kill bacterial growth on skin and mucus membranes Disinfectant: an agent that is used to inhibit/kill bacterial growth on inanimate objects

What factors influence the success of microbial control? What type of organism(s) are you targeting? What type of environment are you treating?

Are all microbes equally sensitive?

What parts of a bacterial cell are sensitive to physical treatments and chemicals? Plasma membrane DNA and proteins

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Physical Methods Heat –Dry: kills by oxidation, ex: incineration –Moist: promotes coagulation of proteins Boiling (100 o C) Autoclave(121 o C, 15lbs/sq inch) Pasteurization –Classic-63 o C for 30 minutes –HTST-72 o C for 15 seconds –UHT-140 o C for 3 seconds

Autoclave

Autoclaves work due to steam under pressure

**Indicators used in autoclaving** What does a color change on the autoclave tape indicate? Why would it be necessary to also place an indicator in the center of the pack? What type of biological indicator is used for quality assurance in an autoclave monitoring program? Why? Where would the biological indicator be placed during the autoclaving procedure?

What if the substance is heat sensitive? Filtration is the best choice Pore sizes can be either.45um or.22um

Physical Methods Low temperature: freezing does not kill bacteria, most cultures are stored at -80 o C Dessication: remove the water and bacteria can remain viable

Physical Methods Radiation –Ionizing radiation (gamma rays, x-rays) –Non-ionizing radiation (uv rays) –Microwaves

Electromagnetic Spectrum

Chemical Methods Most are only able to reduce the numbers of organisms, not achieve sterility Types of chemicals –Phenol and phenolics –Halogens –Alcohols –Heavy metals –Soaps –Quaternary Ammonium compounds –Biguanides As a general rule: if you need to achieve sterility then you should be thinking about physical, not chemical methods of microbial control.

Evaluation of chemicals: disc diffusion method

Chemical control: choosing the right germicidal chemical Goal: sterility vs. disinfection; level of disinfection required dictates potency of chemical required Toxicity: risk benefit analysis Activity in presence of organic material: most are diminished or inactivated Sensitivity of the material to be treated Residue: toxic or corrosive vs residual desired antimicrobial effect Cost and availability Storage and stability: concentrate vs stock solution Environmental risk: antimicrobials in the environment

Chemical control Phenolics Destroy cytoplasmic membranes, denature proteins (binds to tyrosine, alters cell membranes) Triclosan Hexachlorophene Target: vegetative bacteria +/- mycobacteria; not sporocidal; inadequate if viricidal activity is required; leave an active antimicrobial residue

Chemical control Halogens 1. Iodine (binds to tyrosine, alters cell membranes) Tincture Iodophore Target: all bacteria, fungi, most endospores, and some viruses 2. Chlorine (strong oxidizing agent) Target: all types of microbes and viruses; inactivated by organic matter

Chemical Control Alcohols –Mechanism of action is protein denaturation, can also dissolve lipids –Most common are ethanol and isopropanol Target: kill vegetative cells and fungi, do not kill endospores and some naked viruses

Chemical control Heavy metals exert “oligodynamic action” = ability of very low concentrations of metals to kill microbes: interfere with function of enzymes by binding to –SH Silver (and formerly, copper, mercury, lead, arsenic)

Chemical control Surfactants (soaps) –Little value as an antiseptic –Acid-anionic surface-active sanitizers are important in cleaning of dairy equipment and utensils

Chemical control Quaternary Ammonium Compounds (Quats) –Group of positively charged detergents Action: react with the cell membrane → reduced surface tension, membrane damage Target: kill many vegetative bacteria and enveloped viruses, fungi; do NOT kill endospores, Mycobacteria, or naked viruses

Chemical control –Biguanides (chlorhexidine) Used in antiseptic products Adheres to skin and mucus membranes, low toxicity Action: membrane disruption Target: wide range of bacteria, fungi, and some enveloped viruses; does not kill endospores; will not destroy naked viruses

Which antiseptic works the best?